Probiotics and prebiotics are CAM interventions frequently employed by the American public to promote intestinal health and wellness. The presence of beneficial bacteria such as lactobacilli and bifidobacteria, whether delivered exogenously as probiotics or enriched via prebiotics, has been linked to positive health effects including reduction of gut inflammation, diarrhea and allergic reactions. At present, however, our understanding of the mechanism of action underlying these biological effects is significantly lacking. Milk oligosaccharides are naturally evolved prebiotic substrates that facilitate bifidobacterial enrichment and interaction with the host. Work from the UC Davis Milk Bioactives Program has shown that human milk oligosaccharides are utilized by select bifidobacterial strains and demonstrated that small mass oligosaccharide species are uniquely consumed by Bifidobacterium longum subsp. infantis. In addition, genomic analysis of B. infantis isolates has revealed unique gene clusters linked to milk oligosaccharide metabolism that are specifically induced during growth human milk glycans but are not expressed in cells grown on lactose or current commercial prebiotics-inulin or galactooligosaccharides (GOS). There is mounting evidence to indicate that various probiotics interact with intestinal epithelial cells via the production of soluble factors shown to alter intestinal permeability and inhibit inflammatory cascades in epithelial cells. We have obtained preliminary evidence to show a probiotic species, B. infantis, produces a soluble factor(s) that activates gut enteroendocrine (EC) cells. Importantly, growth of B. infantis on milk oligosaccharides markedly increases expression of the soluble factor(s) while growth on lactose, inulin, GOS resulted in little or no production of the factor(s). In addition, growth on milk oligosaccharides also increases the ability of B. infantis to bind to intestinal epithelial cells. Thus, the presence of B. infantis with its preferred growth substrate (milk oligosaccharides) increases the probiotic binding to epithelial cells followed by release of soluble factors capable of activating EC cells. This may result in release of humoral and neuroactive mediators, to initiate changes in GI function, food intake and glucose homeostasis. The overarching aim of this proposal is to characterize the interaction of prebiotic milk oligosaccharides and bifidobacterial species in binding and activation of two types of GI epithelial cells, enterocytes and EC cells and to determine the downstream benefits of this interaction in terms of improved regulation of glucose homeostasis and body weight regulation. We will test the hypothesis that growth of probiotic B. infantis in the presence of milk oligosaccharides results in increased binding to epithelial cells and production of soluble signaling molecules that elicit activation of EC cells and release of regulatory peptides that inhibit food intake and improve glucose homeostasis. This hypothesis will be tested using the following specific aims: 1.To determine the influence of milk oligosaccharides on bifidobacterial binding to intestinal epithelial cells. 2. To determine the mechanism and functional significance of enteroendocrine cell activation by conditioned media from bifidobaceteria growth on MOS. Prebiotics and probiotics play a significant role worldwide as a CAM aimed at intestinal health and wellness. A critical barrier to progress in this field is that the mechanism by which prebiotics and probiotics influence health is ill defined. This proposal will address the role of prebiotic milk oligosaccharides as a mechanism for potentiating probiotic efficacy of B. infantis through increased binding of intestinal cells and production of a soluble signal that activates enteroendocrine cells-the latter of which collectively constitute a diffuse neuroendocrine system of the gut. The significance of this proposal is a greater mechanistic understanding of the beneficial effects of synbiotic (prebioitic plus probiotic) applications and provide clues for treatment of obesity and other disorders that involve the sensory innervation of the gut, such as functional bowel disease.